Methods and devices for cellular transplantation

ABSTRACT

Devices and methods for transplanting cells in a host body are described. The cell comprises a porous scaffold that allows the ingrowth of vascular and connective tissues, a plug or plug system configured for placement within the porous scaffold, and a seal configured to enclose a proximal opening in the porous scaffold. The device may further comprise a cell delivery device for delivering cells into the porous scaffold. The method of cell transplantation comprises a two step process. The device is incubated in the host body to form a vascularized collagen matrix around a plug positioned within the porous scaffold. The plug is then retracted from the porous scaffold, and cells are delivered into the vascularized space created within the porous scaffold.

This application claims priority to U.S. Provisional Application No.61/238,011, filed Aug. 28, 2009, which is incorporated herein byreference in its entirety.

The present disclosure is related to the field of cellular therapy, andmore specifically, to methods and devices for transplantation of cellsinto a host body.

Recent discoveries in the field of cellular therapy present newopportunities for the use of cell transplantation in disease areas withcritical, unmet medical needs. Currently, there are no fully effectivedrug therapies for many acquired and congenital disease conditions, suchas diabetes or Parkinson's disease, which are caused by loss of ordamage to cells producing biomolecules necessary for control ofphysiological functions. Cellular therapy holds the promise of replacinglost or damaged cells with donor cells or stem cells to improve theimpaired physiological functions. For example, transplantation of isletsof Langerhans cells would provide a means of restoring carbohydratecontrol in patients with insulin-dependent diabetes. Similarly,transplantation of dopaminergic neurons or neural stem cells has emergedas a promising cell-based therapy for Parkinson's disease.

Major limiting factors in the application of cellular therapy is thedifficulty in transplanting cells into host tissue and ensuring that thetransplanted cells continue to function without eliciting an immuneresponse or causing other harmful side effects in the host. Attemptshave been made to administer therapeutic cells directly into the hostbody, e.g., in the vascular system or by implantation in an organ ortissue. However, with direct cellular transplantation, the patient isrequired to remain on life-long immunosuppressant therapy, and theimmunosuppressant drugs can cause toxicity to the host and the implantedcells. Additionally, direct exposure of the cells to blood may lead toan immediate blood-mediated inflammatory reaction (IBMIR) that initiatesa coagulation cascade and can destroy a significant portion of thetransplanted cells. Furthermore, cells may become lodged in microvesselsand cause blockage and thrombosis of the vessels, which may result in aloss of function of the transplanted cells and damage to local tissue.

Another therapeutic approach is the delivery of cells using devices thatprovide a biologically suitable environment for the cells to reside inthe host body. Major challenges with this approach are poorincorporation of blood vessels into the device for nourishing the cellsand maintaining an optimal environment within the device for long-termsurvival of the cells. In the absence of an immediately vascularizedenvironment, transplanted cells are not able to obtain enough oxygen oreasily eliminate wastes, and may rapidly die or become damaged throughthe effects of ischemia or hypoxia. Furthermore, even in situationswhere some vessels grow early on, the vessels may not be sustained. Inaddition, the natural inflammatory cascade of the body may also resultin the death of or damage to cells. Some of the other difficultiesencountered with this approach include excessive scarring and/or wallingoff of the device, incompatibility of the device material with thebiological milieu, difficulties in imaging the device and theimplantation environment, improper dimensions of the device affectingbiological function of the cells, inability to load the appropriatenumber of cells for a sustained therapeutic effect, and difficulty inremoving the device when it needs replacement. Furthermore, the deviceconfiguration may not be amenable to the external contours of the body,which can result in abnormal protrusions of the device making the deviceunacceptable to the patient from an aesthetic perspective.

Thus, there still remains a need to find an effective technique forsuccessful transplantation of therapeutic cells. The present disclosureprovides methods and devices for delivering and maintaining cells invivo for an extended period of time, while alleviating many of theproblems associated with existing device-based cell therapy approaches.

In one aspect of the present disclosure, a device for transplantingcells in a host body is provided. The device comprises a porous scaffoldcomprising at least one chamber having a proximal end and a distal end,and at least one removable plug configured to be positioned within theat least one chamber. The porous scaffold comprises a mesh having poressized to facilitate growth of vascular and connective tissues into theat least one chamber. In some embodiments, the porous scaffold comprisesa polypropylene mesh.

Another embodiment of the present disclosure is a device for implantingcells in a host body, wherein the device comprises a porous scaffoldcomprising one or more chambers having a proximal end and a distal end,and an opening at either or both the proximal end and the distal end.The porous scaffold comprises pores sized to facilitate growth ofvascular and connective tissues into the one or more chambers. Thedevice also comprises one or more two-plug systems comprising an outerplug configured to be positioned within the one or more chambers, and aninner plug configured to be positioned within the outer plug.Additionally, the device comprises at least one seal configured toenclose the plug system in the chamber and enclose the opening at eitheror both the proximal end and the distal end of the chamber.

In another aspect of the present disclosure, a method of transplantingcells in a host body is provided. The method comprises the steps ofimplanting a device for holding cells in the host body, wherein thedevice comprises a porous scaffold comprising at least one chamberhaving a proximal end and a distal end. The porous scaffold comprises amesh having pores sized to facilitate growth of vascular and connectivetissues into the at least one chamber. In some embodiments, the porousscaffold comprises a polypropylene mesh. The device further comprises atleast one plug configured to be positioned within the at least onechamber, and the least one chamber comprises an opening at either orboth the proximal end and the distal end. The method comprises the stepsof closing the opening at either or both the proximal end and the distalend of the chamber after implanting the device. The method furthercomprises maintaining the device in the host body until the porousscaffold is infiltrated with vascular and connective tissues, accessingthe device through a surgical incision, reopening either or both theproximal end and the distal end of the chamber, extracting the plug fromthe chamber to create a space within the porous scaffold that isencapsulated in vascularized collagen matrix, delivering a cellpreparation into the vascularized space, and reclosing the opening ateither or both the proximal end and the distal end of the chamber.

In another alternate embodiment, the method of implanting cells in ahost body provides an implantable device for holding cells in the hostbody, wherein the implantable device comprises a porous scaffold havingpores sized to facilitate growth of vascular and connective tissues intothe porous scaffold, at least one two-plug system configured to bepositioned within the porous scaffold. The porous scaffold of theimplantable device comprises at least one chamber having an opening ateither or both a proximal end and a distal end of the chamber. Thedevice comprises a seal to enclose the opening at either or both theproximal and distal ends of the at least one chamber. The at least oneplug system of the implantable device comprises an outer plug configuredto be positioned within the at least one chamber and an inner plugconfigured to be positioned within the outer plug. The method furthercomprises the steps of implanting the device in the host body,maintaining the device in the host body until the device is infiltratedwith vascular and connective tissues, and providing a cell deliverydevice comprising at least one cell infusion tube loaded with a cellpreparation, wherein the cell infusion tube is configured to bepositioned within the outer plug of the at least one plug system.Additionally, the method comprises accessing the implanted devicethrough a surgical incision and opening the seal at either or both theproximal end and the distal end of the device, withdrawing the innerplug from the plug system, inserting the cell infusion tube into theouter plug, withdrawing the outer plug from the at least one chamber andsimultaneously infusing the chamber with the cell preparation, andreclosing the seal. It is to be understood that both the foregoinggeneral description and the following detailed description are exemplaryand explanatory only and are not restrictive of the invention, asclaimed.

Another aspect of the disclosure provides a cellular transplantationdevice comprising a porous scaffold having pores sized to facilitategrowth of vascular and connective tissues into the porous scaffoldcomprising at least one chamber and preferably between 2-12 chambers,wherein the porous scaffold is coated with a biocompatible,biodegradable material designed to temporarily fill the pores of thescaffold. In certain embodiments, the porous scaffold comprises apolypropylene mesh. Suitable biocompatible, biodegradable materialsinclude, e.g., collagen, fibronectin, extracellular matrix proteins, andmembrane cytoskeletal proteins. The disclosure also provides a methodfor transplanting cells into a host body comprising implanting atransplantation device comprising a porous scaffold having pores sizedto facilitate growth of vascular and connective tissues into the porousscaffold comprising at least one chamber and preferably between 2-12chambers, wherein the porous scaffold is coated with a biocompatible,biodegradable material that temporarily fills the pores of the scaffold,and wherein the at least one chamber is filled with the cells to betransplanted and the chamber is sealed.

The accompanying drawings, which are incorporated in and constitute apart of this specification, together with the description, illustratemethods and embodiments of the invention.

BRIEF DESCRIPTION OF DRAWINGS

FIGS. 1A-1E illustrate various embodiments of a single-chamber celltransplantation device consistent with the present disclosure;

FIG. 1F illustrates an embodiment of a multi-chamber celltransplantation device consistent with the present disclosure;

FIGS. 2A-2D illustrate various mesh configurations that may be used forforming a cell transplantation device consistent with the presentdisclosure;

FIG. 3A illustrates a cell transplantation device in accordance with anembodiment of the present disclosure;

FIG. 3B illustrates the components of the cell transplantation device ofFIG. 1A;

FIG. 4 illustrates a porous scaffold of a cell transplantation deviceconsistent with an embodiment of the present disclosure;

FIG. 5A illustrates a seal of a cell transplantation device consistentwith an embodiment of the present disclosure;

FIG. 5B is a cross-sectional view of the seal shown in FIG. 3A;

FIG. 6A illustrates multiple outer plugs of a two-part plug system of acell transplantation device consistent with an embodiment of the presentdisclosure;

FIG. 6B is a cross-sectional view of an outer plug illustrated in FIG.5A;

FIG. 6C is a cross-sectional view of a plug system and a single porousscaffold assembly prior to implantation in a host body;

FIG. 6D is a cross-sectional view of the assembly illustrated in FIG. 4Cfollowing incubation in a host body;

FIG. 6E is a cross-sectional view of a porous scaffold implanted in ahost body following removal of the plug system;

FIG. 7 illustrates multiple inner plugs of a two-part plug system of acell transplantation device consistent with an embodiment of the presentdisclosure;

FIG. 8 illustrates a seal for enclosing cells within a vascularizedchamber of a cell transplantation device consistent with an embodimentof the present disclosure;

FIG. 9A illustrates a device for delivering cells to a celltransplantation device consistent with an embodiment of the presentdisclosure;

FIG. 9B shows a cell infusion mechanism of the delivery deviceillustrated in FIG. 8A;

FIG. 9C shows additional steps of the cell infusion mechanism of thedelivery device illustrated in FIGS. 8A-8B;

FIG. 10 is a flow chart showing the steps of a cell transplantationmethod in accordance with the present disclosure;

FIGS. 11A-11D show a schematic overview of certain steps of a cellinfusion procedure in accordance with the present disclosure;

FIG. 12A shows line graphs of blood glucose measurements afterintraperitoneal implantation of cell transplantation devices, asdescribed in Example 1;

FIG. 12B shows line graphs of blood glucose measurements aftersubcutaneous implantation of cell transplantation devices, as describein Example 1;

FIG. 12C shows line graphs of IVGTT responses in Lewis rats transplantedwith islet cells at 40 days post-transplant, 80 days post-transplant andpost-device removal (at 110 days post-transplant), as described inExample 1;

FIG. 12D shows line graphs of insulin levels in response to glucosechallenge in Lewis rats transplanted with islet cells, as described inExample 1;

FIG. 13A demonstrates histological staining of insulin within thechamber of an implanted device, as described in Example 2;

FIG. 13B demonstrates histological staining of vascularization(microvasculature) within the chamber of an implanted device, asdescribed in Example 2;

FIG. 14 is table of the average collagen thickness and total bloodvessel/cm² calculated for four cell transplantation devices consistentwith embodiments of the present disclosure, as described in Example 3;

FIG. 15A demonstrates tissue incorporation into a cell transplantationdevice at 2, 4 and 8 weeks after implantation, as described in Example3;

FIG. 15B shows blood vessel formation at various margins of an implanteddevice prior to cell transplantation, as described in Example 3;

FIG. 16 shows bar graphs of levels of insulin produced by mature andimmature islets, as described in Example 4;

FIG. 17A demonstrates histological staining of insulin andmicrovasculature within the chamber of an implanted device, as describedin Example 4;

FIG. 17B demonstrates histological staining of microvasculature withinthe chamber of an implanted device after cell transplantation, asdescribed in Example 4;

FIG. 18 shows line graphs of blood glucose levels following isletautograft transplantation, as described in Example 4;

FIG. 19A shows line graphs of absolute blood glucose levels in responseto glucose challenge in Yorkshire-Landrace pigs transplanted with isletcells, as described in Example 4;

FIG. 19B shows bar graphs of Area Under the Curve (AUC) for bloodglucose levels in response to glucose challenge in Yorkshire-Landracepigs transplanted with islet cells, as described in Example 4;

FIG. 19C shows line graphs of fold change in C-peptide levels inresponse to glucose challenge in Yorkshire-Landrace pigs transplantedwith islet cells, as described in Example 4.

DESCRIPTION OF EXEMPLARY EMBODIMENTS

Reference will now be made in detail to embodiments of this disclosure,examples of which are illustrated in the accompanying drawings. Whereverpossible, the same reference numbers will be used throughout thedrawings to refer to the same or like parts. Throughout the disclosure,the terms cell infusion and cell transplantation are usedinterchangeably.

A cell transplantation device for containing therapeutic cells in vivois provided. In one exemplary embodiment, the cell transplantationdevice comprises at least one porous scaffold comprising a chambertherein and having an opening at either or both a proximal end and adistal end of the scaffold, and at least one plug configured to behoused in the chamber. The opening at one or both the ends of thechamber are sized to enable insertion and retraction of the plug fromthe chamber. In one embodiment, the at least one porous scaffold istubular in shape, and the at least one plug is cylindrical and extendsalong a lumen of the at least one porous scaffold. In some embodiments,the porous scaffold is open only at the proximal end. In one suchembodiment, the distal end of the tubular porous scaffold comprises arounded or flat-bottomed surface. In another embodiment, the edges atthe distal end of the porous scaffold are tapered and brought intocontact with one another to seal the distal end.

In another exemplary embodiment, the cell transplantation devicecomprises a porous scaffold comprising one or more chambers having aproximal end and a distal end. The one or more chambers comprise anopening at the proximal end. The device also comprises one or more plugsystems comprising an outer plug configured to be positioned within theone or more chambers, and an inner plug configured to be positionedwithin the outer plug. Additionally, the device comprises at least oneseal configured to enclose the plug system within the chamber and sealthe opening at the proximal end of the chamber.

The porous scaffold is formed of a biocompatible material that shouldelicit only a mild inflammatory response in the body. The mildinflammatory components stimulate angiogenesis and promote incorporationof a vascularized collagen matrix into the device, but do not result insignificant inflammation around the device. An example of such abiocompatible material is polypropylene. In exemplary embodiments, theporous scaffold comprises a woven polypropylene mesh that has sufficientstiffness to facilitate device fabrication. The polypropylene mesh isalso selected to allow microvessels to enter the device and bemaintained as robust, healthy vessels, which is critical for thesurvival and normal functioning of the therapeutic cells infused intothe device.

By encouraging regulated growth of vascularized tissue into the device,the porous scaffold prevents encapsulation of the device with scartissue. Ingrown tissues also stabilize the implant and preventinadvertent movement of the device in situ. Additionally, in someembodiments, the porous scaffold is coated with biological ornon-biological agents to stimulate tissue incorporation andangiogenesis, for example, growth factors. The device may be dip-coatedin a polymer-drug formulation or other known technique to apply thecoating to the device. Examples of biological or non-biological agentsto stimulate tissue incorporation and angiogenesis include but are notlimited to: VEGF (vascular endothelial growth factor), PDGF(platelet-derived growth factor), FGF-1 (fibroblast growth factor),NRP-1 (neuropilin-1), Ang-1, Ang2 (angiopoietin 1,2), TGF-ß, endoglin,MCP-1, αvß3, αvß5, CD-31, VE-cadherin, ephrin, plasminogen activators,angiogenin, Del-1, aFGF (acid fibroblast growth factor), vFGF (basicfibroblast growth factor), follistatin, G-CSF (granulocytecolony-stimulating factor), HGF (hepatocyte growth factor), II-8(interleukin-8), Leptin, midkine, placental growth factor, PD-ECGF(platelet-derived endothelial growth factor), PTN (pleiotrophin),progranulin, proliferin, TGF-α, and TNF-α.

In some embodiments, the outer surface of the porous scaffold isroughened to stimulate tissue ingress. In certain embodiments, theporous scaffold includes various drug-eluting polymer coatings. In otherembodiments, the porous scaffold may be coated with a biodegradable ornon-biodegradable polymer without a drug. The scaffold may be partiallyor completely coated with the polymer. Representative polymers that canbe used for coating and/or drug elution include but are not limited to:methacrylate polymers, polyethylene-imine and dextran sulfate,poly(vinylsiloxane)ecopolymerepolyethyleneimine, phosphorylcholine,poly(ethyl methacrylate), polyurethane, poly(ethylene glycol),poly(lactic-glycolic acid), hydroxyapetite, poly(lactic acid),polyhydroxyvalerte and copolymers, polyhydroxybutyrate and copolymers,polycaprolactone, polydiaxanone, polyanhydrides, polycyanocrylates,poly(amino acids), poly(orthoesters), polyesters, collagen, gelatin,cellulose polymers, chitosans, and alginates or combinations thereof.Additional examples that may be used to coat the scaffold include butare not limited to: collagen, fibronectin, extracellular matrixproteins, vinculin, agar, and agarose. It should be understood thatvarious mixture of the polymers may be used.

With respect to drug elution, in some illustrative embodiments, theporous scaffold includes an antibiotic coating to minimize infections.Representative antibiotics include but are not limited to: ampicillin,tetracycline, nafcillin, oxacillin, cloxacillin, dicloxacillin,flucloxacillin, vancomycin, kanamycin, gentamicin, streptomycin,clindamycin, trimethoprim-sulfamethoxazole, linezolid, teicoplanin,erythromycin, ciprofloxacin, rifampin, penicillin, amoxicillin,sulfonamides, nalidixic acid, norfloxacin, ciprofloxacin, ofloxacin,sparfloxacin, lomefloxacin, fleroxacin, pefloxacin, amifloxacin,5-fluorouracil, chloramphenicol, polymyxin, mitomycin, chloroquin,novobiocin, nitroimadazole. In another embodiment the porous scaffoldincludes a bactericidal agent. Representative bactericidal agentsinclude but are not limited to: benzalkonium chloride, chlorohexidinegluconate, sorbic acid and salt thereof, thimerosal, chlorobutanol,phenethyl alcohol, and p-hydroxybenzoate.

In some other embodiments, parts of the cell transplantation device arecoated with antifibrotic drugs to inhibit fibrous tissue encapsulation.Representative antifibrotic agents include but are not limited to:paclitaxel, everolimus, tacrolimus, rapamycin, halofuginonehydrobromide, combretastatin and analogues and derivatives thereof (suchas combretastatin A-1, A-2, A-3, A-4, A-5, A-6, B-1, B-2, B-3, B-4, D-1,D-2, and combretastatin A-4 phosphate (Oxigene)), docetaxel,vinblastine, vincristine, vincristine sulfate, vindesine, andvinorelbine, camptothecin topotecan, irinotecan, etoposide or teniposideanthramycin, mitoxantrone, menogaril, nogalamycin, aclacinomycin A,olivomycin A, chromomycin A₃, and plicamycin, methotrexate, edatrexate,trimetrexate, raltitrexed, piritrexim, denopterin, tomudex, pteropterin,and derivatives and analogues thereof. In some embodiments, the celltransplantation device may also include polymethyl methacrylate or bonecement or other types of cyanoacrylates.

In some embodiments, the porous scaffold is formed of a material thatallows imaging of the implanted device using, for example, MRIs, fMRIs,CT scans, X-rays, ultrasounds, PET scans, etc. In one such embodiment,the porous scaffold comprises a polymer mesh (for example,polypropylene, polytetrafluoroethylene (PTFE), polyurethane, polyesters,silk meshes, etc.) that is immunologically compatible and allows imagingof the neovascularized tissue. In another embodiment, the porousscaffold comprises a combination of materials. In one such embodiment,the porous scaffold comprises interwoven polypropylene and silk strands.

The pore size of the scaffold material is selected to facilitate tissueincorporation and vascularization within the chamber of the porousscaffold. In some embodiments, the pore sizes may range from about 50 nmto 5 mm. In one exemplary embodiment, the porous scaffold comprises awoven polypropylene mesh with 0.53 mm pore diameter.

In some embodiments, the pore size is selected to exclude immune cellsor immune agents from penetrating the implanted device. In some otherembodiments, the pore size does not necessarily need to exclude immunecells or immune agents from infiltrating the device. This would be thecase, for example, when the device is used to transplant a combinationof cells, including immunoprotective cells, (e.g. Sertoli cells,mesenchymal stem cells, etc.) which can provide immune protection to theco-transplanted cells. This would also be the case, for example, whenthe device is used to transplant syngeneic cells, or cells derived fromthe patient receiving the transplant.

The plug or plug system of the cell transplantation device is configuredto fit into the chamber within the porous scaffold. The plug or plugsystem may comprise a non-porous material (e.g., polytetrafluoroethylene(PTFE), polypropylene, etc.) that inhibits ingrowth of biological tissueinto the plug or plug system. The plug or plug system may be a hollow orsolid structure. However, if a hollow plug is used, care should be takento prevent infiltration of collagen or any other biological materialinto the lumen of the plug when the device is implanted into hosttissue. The plug system is discussed in further detail below.

In some embodiments, the proximal end of the plug or plug system isconnected to a seal. In such embodiment, the seal is configured to closethe proximal opening of the chamber when the plug or plug system iscompletely inserted into the chamber of the porous scaffold. The seal isstructured to hold the plug or plug system in place inside the porousscaffold. In another embodiment, the seal is separate from the plug orplug system. In yet another embodiment, the seal is connected to theporous scaffold. Further, in some exemplary embodiments, the proximalend of the chamber is closed using surgical sutures and/or vascularclips without using a separate seal.

When implanted in a host body, the porous scaffold of the deviceencourages ingrowth of vascular and connective tissue, such that theplug or plug system housed within the scaffold becomes encapsulated in avascularized tissue matrix. When the plug or plug system is removed fromthe porous scaffold, a neovascularized chamber is created within thedevice, which can then be used for holding a cell preparation in thehost body.

The sizes of the porous scaffold and the plug or plug system areselected to provide an optimal surface area-to-volume ratio for holdingcells in vivo and for ensuring long-term survival of the cells withinthe neovascularized chamber. Similarly, the number of chambers in thetransplantation device is determined based on the volume and/or numberof cells that are to be transplanted. In some embodiments, the totalvolume of the cell transplantation device is adjusted by increasing ordecreasing the number of chambers while maintaining an optimum surfacearea-to-volume ratio of each individual chamber. In other embodimentsthe length of the chambers is adjusted to alter the total volume.Alternatively, in various embodiments, the cell transplantation devicecomprises a fixed number of chambers, but only a selected number ofchambers are infused with cells depending on the total volumerequirement of the device. In other embodiments the length of thechambers is adjusted as well as the number of chambers to alter thetotal volume required.

The cell transplantation device disclosed can be implanted eithersubcutaneously or intraperitoneally in a host body, including theomentum or other appropriate site. Alternatively, the cell implantationdevice disclosed can be implanted partially intraperitoneally in a hostbody, including into the omentum or other appropriate site and extendinto the subcutaneous environment. In one embodiment the cells may beloaded in the portion of the device extending into the subcutaneousenvironment while the rest of the device is in the intraperitonealenvironment. In another embodiment the cell transplantation device maybe implanted into the brain, spinal cord area or any other organ asrequired to elicit a therapeutic effect from transplanted cells. In mostinstances, the host is a human, but may be another mammal ornon-mammalian animal. The cell transplantation procedure is a two-stepprocess comprising a device implantation step followed by a cellinfusion (cell transplantation) step. The cell infusion step isimplemented after an in vivo incubation period during which theimplanted device is infiltrated with a vascularized collagen matrix. Inone embodiment, the incubation period is approximately thirty days,which allows adequate time for angiogenesis and collagen infiltration ofthe porous scaffold. The incubation period may be lengthened orshortened, depending on the degree of neovascularization and tissue(collagen with cells) formation needed or desired. For example,transplantation devices may vascularize at different rates depending onthe device material, dimensions, or coatings, such as, for example,antibiotic coatings, growth factors, etc. Transplantation devices mayalso vascularize at different rates in different hosts, or when locatedin different body tissues within the same host. It is within the skillof a person in the art to determine the appropriate incubation period.For example, imaging studies may be performed prior to delivering cellsto ensure that adequate vascular and/or connective tissue is depositedaround and through the walls of the porous scaffold during theincubation period. For the cell infusion step, the implantation site isaccessed through a surgical incision, and the plug or plug system isremoved from the porous scaffold to create a collagen and blood vessellined pocket within the scaffold. The cell preparation is then deliveredinto the vascularized pocket, and the porous scaffold is re-sealed. Inanother embodiment the cell transplantation procedure is a single stepprocess whereby the device is placed and the cells implanted at the sametime. In this circumstance, the cells may be placed in a matrix so thatthey do not leak through the pores of the device or alternatively thedevice may be coated with a degradable polymer to prevent cells fromleaking from the device during the process of collagen and angiogenesisdevelopment.

In some embodiments, the cells to be transplanted may be combined with abiocompatible viscous solution or biodegradable polymer formulationprior to being loaded into the chamber of any of the transplantationdevices described herein. This biodegradable polymer will protect thecells until the device is fully vascularized by the host body. Theseformulations may be placed in the chambers prior to or followingplacement of the device in a host, but before a collagen matrix andvascular structures have formed in the device. Cells combined with abiocompatible viscous solution or biodegradable polymer formulation willbe particularly useful in devices designed to be loaded with cells priorto implantation of the device in the host body. Representative polymersthat can be used as a biodegradable formulation in conjunction withcells include but are not limited to: polyethylene-imine and dextransulfate, poly(vinylsiloxane)ecopolymerepolyethyleneimine,phosphorylcholine, poly(ethylene glycol), poly(lactic-glycolic acid),poly(lactic acid), polyhydroxyvalerte and copolymers,polyhydroxybutyrate and copolymers, polydiaxanone, polyanhydrides,poly(amino acids), poly(orthoesters), polyesters, collagen, gelatin,cellulose polymers, chitosans, alginates, fibronectin, extracellularmatrix proteins, vinculin, agar, agarose, hyaluronic acid, matrigel andcombinations thereof.

It should be noted that cells may be placed in the device; however, thecells may also be encapsulated. The following are by way of example andnot by way of limitation. Examples of polymeric cell encapsulationsystems include alginate encapsulating, polysaccharide hydrogels,chitosan, calcium or barium alginate, a layered matrix of altinate andpolylysine, photopolymerizable poly(ethylene glycol) polymer toencapsulate individual cells or cell clusters, polyacrylates includinghydroxyethyl methacrylate methyl methacrylate, silicon capsules, siliconnanocapsules, and polymembrane (acrylonitrile-co-vinyl chloride).

FIGS. 1A-1E illustrate various exemplary embodiments of a celltransplantation device 1. Device 1 comprises a polymer mesh (e.g. apolypropylene mesh, a PTFE mesh, or any other suitable material) thatforms a porous chamber 2 for containing cells in a host body. In someembodiments, device 1 may comprise 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12or more porous chambers 2. The availability of multiple chambers allowsthe use of any number or combination of chambers depending on the volumeof cellular preparation required, which is within the knowledge andskill of persons skilled in the art to determine.

As shown in FIG. 1A, device 1 comprises a proximal end 3, a distal end4, and a plug 5 housed in porous chamber 2. In one embodiment, porouschamber 2 is tubular in shape, and plug 5 is cylindrical and extendsalong a lumen of porous chamber 2. In another exemplary embodiment,porous chamber 2 comprises an opening at proximal end 3. The opening atproximal end 3 is sized to enable insertion and retraction of plug 5from porous chamber 2. In one such embodiment, the opening at proximalend 3 is sealed using surgical sutures and/or vascular clamps duringdevice incubation and after infusion of cells into the device. As wouldbe understood by a person of ordinary skill in the art, any othersurgical sealing element, for example, microvascular clips, clamps,etc., can be used to seal the opening at proximal end 3. In anotherembodiment, device 1 comprises a non-porous flap 6 at proximal end 3, asillustrated in FIG. 1B. In one such embodiment, flap 6 is made ofsilicone. Flap 6 can be sealed using surgical sutures, clamps or anyother suitable sealing mechanism during device incubation and afterinfusion of cells into the device. In an exemplary embodiment, distalend 4 of device 1 comprises a rounded or flat-bottomed surface. Inanother embodiment, device 1 comprises an opening at distal end 4, whichcan be sealed using surgical sutures, clamps or any other surgicalsealing element, during device incubation and after infusion of cells.In yet another exemplary embodiment, as shown in FIG. 1C, distal end 4comprises a non-porous portion 7, which prevents tissue ingrowth at thedistal end of the device and facilitates retraction of plug 5 from thedevice prior to cell infusion.

In some embodiments, as illustrated in FIG. 1D, the proximal end of plug5 is connected to a seal 8. In such an embodiment, seal 8 is configuredto close the opening at proximal end 3 when plug 5 is inserted intochamber 5. Seal 8 is structured to hold plug 5 in place inside theporous chamber. In another embodiment, plug 5 is longer than porouschamber 2 and acts as a seal on both proximal end 3 and distal end 4 ofthe device, as shown in FIG. 1E. The edges of porous chamber 2 aroundplug 5 are sealed using surgical sutures and/or surgical glue. Afterremoval of plug 5 prior to cell infusion, the openings at proximal end 3and distal end 4 can be sealed using surgical sutures, vascular clamps,or any other suitable sealing mechanism, as would be understood by oneof ordinary skill in the art.

In some exemplary embodiments, device 1 comprises multiple porouschambers 2 that are laterally connected to each other. In one suchembodiment, the multiple porous chambers 2 are formed, for example, byultrasonically welding the top and bottom surfaces of a porous materialalong a line substantially parallel to a longitudinal axis of thedevice. FIG. 1F illustrates a cell transplantation device having eightporous chambers 2. Each chamber 2 houses a plug 5 during the deviceincubation phase. Plugs 5 are removed from chambers 2 prior to infusionof cells into the chambers. In one embodiment, device 1 comprises eightporous chambers and has an overall length of 50 mm and width of 45 mm.Each porous chamber 2 has an inner diameter no greater than 3.5 mm andhouses a plug 5 having a length of approximately 40 mm and diameter 2.5mm. In such an embodiment, plug 5 is formed of a non-porous,biocompatible material, for example, polytetrafluoroethylene (PTFE).

Exemplary embodiments of the cell transplantation device of the presentdisclosure are formed of medical grade polypropylene meshes, forexample, Polypropylene Knitted Mesh (PPKM) purchased from SURGICALMESH™,Brookfield, Connecticut, USA. In illustrative embodiments, the meshesare formed of monofilaments ranging in diameter from 0.1 mm to 0.3 mm,and mesh pore sizes ranging from 0.3 mm to 1 mm, from 0.4 mm to 0.85 mmand 0.5 mm to 0.6 mm. FIGS. 2A-2D illustrate various exemplary meshconfigurations that can be used for forming the cell transplantationdevices. FIG. 2A illustrates a polypropylene mesh (PPKM601) having apore size of 0.5 mm and monofilament thickness of 0.3 mm; FIG. 2B showsa polypropylene mesh (PPKM602) having a pore size of 0.53 mm andmonofilament thickness of 0.18 mm; FIG. 2C shows a polypropylene mesh(PPKM404) having a pore size of 0.53 mm and monofilament thickness of0.13 mm; and FIG. 2D shows a polypropylene mesh (PPKM604) having a poresize of 0.85 mm and monofilament thickness of 0.2 mm.

FIG. 3A illustrates another exemplary embodiment of a celltransplantation device 10. FIG. 3B illustrates the components of thecell transplantation device 10. Device 10 comprises a porous scaffold12, a primary seal 14, at least one plug system comprising an outer plug16 and an inner plug 18, and a secondary seal 20.

As illustrated in FIG. 4 , porous scaffold 12 of cell transplantationdevice 10 may comprise a polymer mesh (e.g. a polypropylene mesh, a PTFEmesh, or any other suitable material) that forms one or more porouschambers 22 for containing cells in a host body. In some embodiments,the porous scaffold 12 may comprise 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11,12 or more porous chambers 22. The availability of multiple chambersallows the use of any number or combination of chambers depending on thevolume of cellular preparation required, which is within the knowledgeand skill of persons skilled in the art to determine.

Porous chambers 22 may be created, for example, by joining the top andbottom surfaces of porous scaffold 12 along a line substantiallyparallel to a longitudinal axis of the device. Multiple porous chambers22 may have equal or different cross-sectional dimensions and surfaceareas. In one embodiment, multiple porous chambers 22 are formed byultrasonically welding the polymer mesh from a proximal end 24 to adistal end 26 of the scaffold. The top and bottom surfaces of porousscaffold 12 are continuous across the one or more porous chambers 22,interrupted only by ultrasonic weld lines 28, which run substantiallyparallel to a longitudinal axis of porous scaffold 12. The top andbottom surfaces of porous scaffold 12 can be indented slightly at eachweld line, which offers additional surface area for vascularization andprovides physical stability to device 10 within a host. In oneembodiment, the edges at distal end 26 are tapered and ultrasonicallywelded to one another to seal the distal end 26.

With reference to FIG. 3B, primary seal 14 is configured to seal the oneor more porous chambers 22 during device incubation and after cellinfusion. Primary seal 14 comprises an inert and biocompatible polymericfilm or any other suitable material. In one embodiment, primary seal 14is ultrasonically welded at the lateral edges and at the taperedproximal end 31, as illustrated in FIGS. 5A and 5B. Distal end 32 ofprimary seal 14 is attached to proximal end 24 of porous scaffold 12. Inone embodiment, distal end 32 is ultrasonically welded to proximal end24 of porous scaffold 12.

In various embodiments, primary seal 14 comprises a re-sealable lock 34,which assists in maintaining the at least one outer plug 16 within aporous chamber 22 during the incubation period. Lock 34 also preventsleakage of cellular preparation during the cell infusion process. Anysuitable re-sealable locking mechanism may be used as lock 34. In oneembodiment, lock 34 comprises interlocking groove and ridge features,which form a tight seal when pressed together and unlocks when the topand bottom surfaces of seal 14 are pulled apart at the proximal end 31.Following the device incubation period, access to outer plug 16 isachieved by trimming proximal end 31 of primary seal 14 and openingre-sealable lock 34. After the cell preparation is delivered into porousscaffold 12, lock 34 is reclosed and proximal end 31 is re-sealed using,e.g., surgical sutures, staples or bioadhesives, or hermetic seals.

The number of plug systems may correspond to the number of porouschambers 22 in cell transplantation device 10. Outer plug 16 is housedwithin porous chamber 22 during the device incubation period. In someembodiments, the length of outer plug 16 is approximately equal to thelength of the respective porous chamber 22. As illustrated in FIG. 6A,in one embodiment, multiple outer plugs 16 are connected at a proximalend 40 using a common spine 42. Common spine 42 may include one or moregrooves 43 to facilitate removal of outer plugs 16 from porous chambers22. For example, grooves 43 may allow common spine 42 to be graspedusing forceps.

In some embodiments, outer plug 16 has a hollow core 45 that houses aninner plug 18. As shown in FIG. 6B, in one embodiment, hollow core 45 isconstrained with one or more internal bosses 47 along the length of theinner surface of the plug. Internal bosses 47 provide an air spacebetween the outer plug 16 and the inner plug 18, which allows trappedair bubbles to escape during the delivery of the cellular preparation,which is described in further detail below. The air space also preventsvacuum formation during the removal of inner plug 18, and thereby,maintains the integrity of the newly formed vascularized collagen matrixin and around the porous chamber. Thus, in some aspects, the plug systemcomprising outer plug 16 and inner plug 18 may facilitate delivery ofcells to the cell transplantation device 10, and may also increase thechances of cell survival within an intact collagen matrix.

In some embodiments, proximal end 40 and distal end 41 of outer plug 16comprise sealing mechanisms, for example, internal grooves or taperedsurfaces, to ensure an effective seal with inner plug 18. As shown inFIG. 7 , proximal end 50 and distal end 51 of inner plug 18 may includecomplementary sealing mechanisms 53 to prevent infiltration of collagenmatrix into hollow core 45 during the incubation period. For example, inone embodiment, sealing mechanism 53 comprises a groove extending aroundthe periphery of the proximal and distal ends of inner plug 18, andouter plug 16 comprises a ridge around the periphery of its distal andproximal ends. In such an embodiment, the ridge on outer plug 16 and thegroove on inner plug 18 interlock when inner plug 18 is inserted intothe hollow core 45 of outer plug 16, so as to form a complete sealbetween the inner and outer plugs and prevent permeation of anybiological material into hollow core 45. Additionally, in suchembodiments, if outer plug 16 comprises one or more internal bosses 47,the height of the ridges at the proximal and distal ends of outer plug16 may be greater than the height of the internal bosses 47.

FIGS. 6C and 6D illustrate cross-sectional views of porous chamber 22and plug 16, 18 assembly, in accordance with one embodiment of thepresent disclosure. FIG. 6C is a cross-sectional view of the assemblyprior to implantation in a host body, and FIG. 6D illustrates thecross-sectional view of the assembly after incubation in a host body.The inner diameter of porous chamber 22 and outer diameter of outer plug16 are selected to maintain a space 46 around the periphery of outerplug 16 for tissue formation. For example, in one illustrativeembodiment, the inner diameter of porous chamber 22 is no greater than4.5 mm and the outer diameter of plug 16 is no greater than 3.5 mm. Inanother embodiment, the inner diameter of porous chamber 22 is nogreater than 3.5 mm and the outer diameter of plug 16 is no greater than2.5 mm. These embodiments provide, for example, approximately 0.5 mm ofspace around outer plug 16 for formation of a vascularized collagenmatrix. The space around outer plug 16 also offers sufficient room forinsertion and retraction of the outer plug into and out of the porouschamber.

When cell transplantation device 10 is implanted in a host body,vascular and connective tissues penetrate through porous chamber 22 intospace 46 and form a vascularized tissue matrix 48 around outer plug 16.Plug 16 prevents penetration of tissue matrix 48 further into the lumenof porous chamber 22. When inner plug 18 and outer plug 16 are retractedfrom porous chamber 22, a pocket 49 is created within porous chamber 22,which may be used for containing cells in the host body. Pocket 49 isenveloped in vascularized tissue matrix 48, as shown in FIG. 6E.

The number of inner plugs 18 may correspond to the number of outer plugs16. Inner plug 18 is housed within hollow core 45 of outer plug 16during the device incubation phase. In one embodiment, multiple innerplugs 18 are connected at a proximal end 50 using a common spine 52. Insome embodiments, common spine 52 comprises a clip feature 54 to assistin the handling of inner plug 18 during extraction from outer plug 16.

Secondary seal 20, as illustrated in FIG. 8 , is used to contain thecellular preparation in the porous chambers when the primary seal 14 isreclosed after delivery of a cell preparation into the celltransplantation device 10. Secondary seal 20 is positioned at proximalend 24 of porous scaffold 12 after the cell preparation is completelydelivered into porous chamber 22 and outer plug 16 is retracted fromdevice 10. In some embodiments, secondary seal 20 comprises grooves 60to facilitate insertion into device 10 using tweezers.

In another aspect of the present disclosure, a device and method fordelivering cells into a cell transplantation device are disclosed, andwill be explained with reference to cell transplantation device 10. FIG.9A illustrates the various components of a cell delivery device 70. Thecell delivery device 70 comprises at least one cell infusion tube 71,connector cap 72 having a clip feature 73, and connector spacer 74.

Cell infusion tube 71 may comprise polymeric tubing (e.g. polyethylenetubing) or any other suitable material to deliver the cell preparationinto porous chamber 22 of device 10 during the cell infusion step. Thenumber of cell infusion tubes in the delivery system may correspond tothe number of porous chambers 22.

Connector spacer 74 is positioned at the distal end of cell infusiontube 71 and couples or interfaces with the proximal end 40 of outer plug16 during the cell delivery process. Connector spacer 74 includes one ormore through-holes through which cell infusion tube 71 is inserted, asshown in FIG. 9A. The through-holes are configured to provide a lightinterference fit with cell infusion tube 71. The fitting is adapted tokeep cell infusion tube 71 in place during the cell infusion process.Additionally, in certain embodiments, connector spacer 74 comprisesvents 76 to expel air from the air spaces in outer plug 16 created byinternal boss 47 during the cell delivery process, as described furtherbelow. In one embodiment, outer plug 16 comprises a hub 78 at theproximal end 40. In such an embodiment, connector spacer 74 is insertedinto hub 78 during the cell infusion process to secure the deliverydevice 70 to the cell transplantation device 10.

The proximal end of cell infusion tube 71 comprises connector cap 72. Asthe tube is inserted into outer plug 16, connector cap 72 advancesdistally towards connector spacer 74. When tube 71 is completelyinserted into outer plug 16, connector cap 72 fits over connector spacer74 and/or hub 78, and clip feature 73 connects with outer plug 16/or hub78 along common spine 42, as shown in FIG. 9C. This enables connectorcap 72, connector spacer 74, and outer plug 16 to be retracted as asingle unit as the cell preparation is infused into porous chamber 22.

In yet another aspect of the present disclosure, a method for cellulartransplantation is disclosed and will be explained with reference tocell transplantation device 10 and cell delivery device 70. The celltransplantation method is not limited to the device embodimentsdisclosed herein and may be used with any cell transplantation and celldelivery devices.

FIG. 10 is a flowchart illustrating the steps of an exemplary celltransplantation procedure. The cell transplantation procedure isgenerally a two-step process comprising a device implantation stepfollowed by a cell infusion step. Device 10 is implanted in the hostbody prior to delivery of cells to allow adequate time for collagen andblood vessels to infiltrate porous scaffold 12. In some embodiments,device 10 is sterilized using ethylene oxide prior to implantation. Thedevice 10 may be packaged in a self-seal package or any othersterilizable package along with a sterility indicator strip for anethylene oxide-based sterilization process. In some other embodiments,gamma radiation or dry heat autoclaving is used to sterilize the deviceprior to implantation. The type of sterilization method used isdependent on the scaffold material, since dry heat autoclaving is knownto warp certain polymeric materials (e.g. polypropylene) due to low heatdeflection temperature. Gamma radiation, at a sterilization dose of 6M-Rad, can successfully sterilize cell implantation devices; however,gamma radiation may decrease the shelf life of devices made ofpolypropylene.

Device 10 may be implanted subcutaneously or intraperitoneally. Forexample, for subcutaneous implantation of the device in the host body,an incision is made through the dermis and epidermis followed by carefulblunt dissection of connective tissue and adipose, creating asubcutaneous pocket caudal to the incision line (step 810). Once anadequate space is created (roughly the dimensions of the device), device10 is implanted into the subcutaneous pocket, and the incision issutured (step 820). Alternatively, device 10 may be implanted in theperitoneal cavity through an abdominal incision. The device implantationsteps (steps 810 and 820) are followed by a device incubation period(step 830) during which a vascularized collagen matrix is deposited inand around porous scaffold 12.

After the incubation period, device 10 is accessed through a secondsurgical incision. For example, proximal end 31 of primary seal 12 maybe trimmed in situ to open device 10 (step 840). Inner plug 18 is thenextracted from outer plug 16 and discarded (step 850). During the innerplug removal process, air movement is facilitated by internal bosses 47,which prevent formation of a vacuum inside the device, which can causedisruption of any newly formed blood vessels in and around the device.Removal of inner plug 18 disengages proximal end 50 and distal end 51 ofinner plug 18 from proximal end 40 and distal end 41 of outer plug 16. Acellular preparation is then delivered into device 10 using celldelivery device 70.

FIGS. 11A-11D show a schematic overview of certain steps of an exemplarycell infusion procedure and will be explained with reference to theflowchart shown in FIG. 10 . For administering the cells into device 10,cell infusion tube 71 of delivery device 70 is loaded with cellularpreparation 79, and the tube is inserted into the hollow core 45 ofouter plug 16, as shown in FIG. 11A (step 860). Connector spacer 74couples with the proximal end 41 and/or hub 78 of outer plug 16. As tube71 is advanced into the outer plug, air is vented through internalbosses 47 of outer plug 16 and vents 76 of connector spacer 74. Whentube 71 is advanced all the way into outer plug 16, connector cap 72interfaces with connector spacer 74. Clip 73 of connector cap 72 is thenconnected to hub 78 of outer plug 16 (step 870). In this case, outerplug 16, connector cap 72 and connector spacer 74 are then retractedslightly from porous chamber 22 as a single unit to create a space atthe distal end of porous chamber 22 (step 875). In some embodiments,outer plug 16 may be retracted slightly from porous chamber 22 prior toconnecting delivery device 70 with outer plug 16. In other words, step875 may be performed prior to step 870. Gentle pressure is applied to asyringe connected to cell infusion tube 71 to deliver the cells intoporous chamber 22 (step 880). Care is taken to ensure tube 71 remains inthe porous chamber 22 as pressure is applied to deliver the cellularpreparation.

In one embodiment, outer plug 16 is retracted approximately 5 mm beforethe cell infusion is started, as illustrated in FIG. 11B. As pressure(P) is applied to the syringe connected to cell infusion tube 71, thecell preparation 79 infuses into the porous chamber 22. As the cellpreparation is delivered into porous chamber 22, outer plug 16 and cellinfusion tube 71 are withdrawn from the device, as shown in FIGS. 11Cand 11D (step 885). When the device is completely filled with thecellular preparation 79, cell infusion is stopped and cell infusion tube71 is completely retracted from device 10 (step 890). Porous chamber 22is then evaluated for remaining capacity for cellular preparation, andany remaining cell preparation may be carefully added to the end of theporous chamber. The cell preparation is contained within the porouschamber 22 by placing secondary seal 20 at the proximal end 40 of porouschamber 22, followed by closing the re-sealable lock 34 of primary seal12, and securing the proximal end 31 of primary seal 12 with surgicalsutures or staples or other suitable sealing mechanisms (step 895).Finally, the surgical incision is closed using surgical sutures, staplesor tissue adhesives, thereby completing the cell transplantationprocedure.

The devices and methods for cell transplantation disclosed can be usedfor transplantation of any therapeutic cells, or a combination of cells,into a host body for providing therapeutic biological material to thehost for the treatment of a disease condition. The cells may beallogeneic, xenogeneic or syngeneic donor cells, patient-derived cells,including stem cells, cord blood cells and embryonic stem cells. Thestem cells may be differentiated into the appropriate therapeutic cells.The cells may be immature or partially differentiated or fullydifferentiated and mature cells when placed into the device. The cellsmay also be genetically engineered cells or cell lines. In one aspect,an embodiment consistent with the present disclosure is used fortransplantation of islets of Langerhans cells to provide means for bloodglucose regulation in the host body. In another aspect, an embodiment ofa cell transplantation device is used for co-transplantation of isletsof Langerhans and Sertoli cells, where the Sertoli cells provideimmunological protection to the islet cells in the host body. The immuneprotection provided by Sertoli cells in a host body was previouslydisclosed, for example, in U.S. Pat. No. 5,725,854, which isincorporated herein by reference in its entirety. Accordingly, thisdisclosure also contemplates methods of treating various diseases bytransplanting therapeutic amounts of cells to subjects in need thereofusing an embodiment of a cell transplantation device as disclosed here.

The density of the transplanted therapeutic cells, or combinations ofcells, is determined based on the body weight of the host and thetherapeutic effects of the cells. As noted earlier, the dimensions ofthe cell transplantation device and number of porous chambers to be used(in a multi-chamber device) are determined based on the number of thecells required, the extent of vascularization achievable during thedevice incubation period, and the diffusion characteristics of nutrientsand cellular products in and out of the implanted devices.

Examples

The following examples are provided to better explain the variousembodiments and should not be interpreted in any way to limit the scopeof the present disclosure. The cell transplantation devices used inthese examples are formed of polypropylene meshes and comprise a singlePTFE plug in each porous chamber of the devices.

1. Cell Transplantation Devices Containing Islet Cells are Capable ofRestoring Normoglycemia in Lewis Rats

Cell transplantation devices were used for implanting syngeneic isletcells in Lewis rats for restoring normoglycemia. The glucose response ofthe implanted cells was compared with the glucose response of isletcells administered directly into the portal veins of rats. The Lewisrats were divided into three study groups, with nine rats in each group.In the first and second study groups, the devices were implanted inintraperitoneal and subcutaneous cavities, respectively. In the thirdgroup, the islet cells were administered directly into the portal veins.

The implanted devices were incubated in the Lewis rats for at least onemonth to allow vascular ingrowth. Diabetes was then chemically inducedin the rats by injecting streptozotocin. The rats were considereddiabetic if three successive blood glucose readings were at least 18.0mM. Isolated Lewis rat islet cells (10,000 IEQ/Kg weight) were theninfused into the implanted devices or directly into the portal veins ofdiabetic rats. Insulin pellets were removed at 14 days post islettransplantation (denoted by the closed rectangle above the graphs inFIGS. 11A and 11B). Blood glucose levels in the rats were monitored fora period of 100 days. At 100-days post-transplantation, the devices wereremoved to confirm that the transplanted islets were responsible forreversal of diabetes.

FIGS. 12A and 12B show glucose normalization results for rats receivingintraperitoneal (omental chamber) and subcutaneous cell transplantationdevices, respectively. Successful cell transplantation resulted innormalization of blood glucose levels (glucose reading less than 8.0mM), as denoted by the solid traces. The transplants not achievingnormoglycemia are denoted by dotted traces. The results indicate thatnormal glycemic level was maintained in a statistically significantnumber of diabetic rats that received the islet cells. Following theremoval of the implanted devices at 100-days post-transplantation, ratswhich previously demonstrated normal glycemic levels returned tohyperglycemic levels, indicating that the devices contained fullyfunctioning grafts that were responsible for achieving normoglycemiaprior to device removal. The rate at which blood glucose concentrationsreached non-diabetic levels was statistically different between thestudy groups (p<0.0001, t-test).

FIG. 12C shows IVGTT (intravenous glucose tolerance test) responses inLewis rats transplanted with islet cells. IVGTTs was performed at 40days and 80 days post-transplantation The glucose response of rats withintraperitoneal and subcutaneous transplants were compared againstglucose response of rats that received intra-portal islet cells. IVGTTswere performed on three rats in each study category. At 40-days and80-days post transplantation, the blood glucose levels in ratstransplanted with islet cells dropped below 8.0 mM within 50 minutes ofreceiving a glucose challenge, as shown in FIG. 12C. The celltransplantation devices were removed at 100-days. The blood glucoselevels did not drop when a bolus of glucose was administered at110-days, indicating that the transplanted islet cells were responsiblefor the normoglycemia achieved in the diabetic rats prior to removal ofthe implanted devices.

FIG. 12D show insulin responses in Lewis rats transplanted with isletcells. The insulin levels were tested using enzyme linked immunosorbentassays (ELISA). The analysis was performed in triplicate. The resultsindicate a significant difference in blood insulin levels upon glucosechallenge (p<0.005, t-test). As shown in FIG. 12D, the insulin levels inrats that received the transplanted devices correlated well with theinsulin levels in rats that received intra-portal islet cells.

2. Histological Detection of Insulin and Vascularization within thePorous Chambers of Cell Transplantation Devices

Following removal of the implanted devices at 100-days, insulin wasdetected in the devices using specific primary antibodies againstinsulin. FIG. 13A shows the result of the insulin staining within theporous chamber of a subcutaneously implanted device. The detection ofinsulin within the chamber indicated that the islet cells contained inthe devices were viable and functional at 100-days post-transplantation.

Histological evaluation of implanted devices was also performed toverify the formation of vascular tissue in the collagen matrix depositedin and around the devices. Immunohistochemical staining for Factor VIIIassociated with endothelial cells indicated well-formed vascularstructures deeply embedded in connective tissue, as shown in FIG. 13B(dark structure indicates endothelium; cell nuclei are indicated byarrows). The histological evaluation also demonstrated the penetrationof neovascularized tissue towards the core of the cell transplantationdevices.

3. Assessment of Angiogenesis and Collagen Deposition in CellTransplantation Devices

To determine the appropriate length of the implantation phase (timebetween implantation of device and engraftment of islets), celltransplantation devices were implanted subcutaneously into eight weekold Yorkshire-Landrace pigs for 2, 4 and 8 weeks. Following implantationfor the respective time period, the devices were explanted and analyzedto determine the level of angiogenesis and collagen deposition.

a) Gross Assessment of Angiogenesis and Collagen Deposition

Photographs were taken of both the ventral and dorsal surfaces of theexplanted devices for gross analysis of blood vessel and tissueformation. A 1 cm×1 cm grid was laid over the photographs to quantifythe microvessel and tissue (collagen with cells) formation. Each 1 cm²box within the grid was scored for vessel formation, allowing for atotal vessel/cm² to be calculated for the entire surface of theexplanted devices. The average thickness on the medial and lateralperimeters of the devices were measured to evaluate the amount ofcollagen deposition. FIG. 14 shows a table of the average collagenthickness and total blood vessel/cm² calculated for four devices formedusing different porous materials (meshes). Sufficient microvessel andtissue formation was observed for all the four mesh types at 2 weekspost-implantation. The results also indicate that the amount of timerequired for microvessel formation and collagen deposition may varydepending on the device material (porosity, surface roughness, etc. ofthe meshes).

b) Histological Analysis of Angiogenesis and Collagen Deposition

Angiogenesis was determined by staining endothelial cells withHematoxylin and Eosin (H&E) stain (FIG. 15A) and von Willebrand factor(FIG. 15B). FIG. 15A demonstrates tissue incorporation into the devicesat 2, 4 and 8 weeks after implantation. FIG. 15B shows blood vesselformation at various margins of a device prior to cell transplantation.The assessment of tissue incorporation into the devices showed that thedevices incorporate collagen and microvessels at all measured timepoints prior to islet transplantation.

4. Assessment of Cell Transplantation Devices Receiving PorcineAutograft Islets

Eight week old Yorkshire-Landrace pigs were implanted with celltransplantation devices for four and eight weeks. To make the animalsdiabetic, a 90% pancreatectomy was performed followed by a 150 mg/Kgintravenous dose of streptozotocin one day after the surgery. Isletswere isolated from the pancreas before performing the pancreatectomy.The immature islet grafts were transplanted into the animals five daysafter graft isolation and pancreatectomy to allow sufficient time forrecovery and confirmation of diabetes.

The insulin producing capacities of the immature islet cells were testedprior to transplantation. As shown in FIG. 16 , the immature isletsproduced about 10% of the insulin normally expected from adult islets.This fact combined with the low islet transplant number of about 3-5KIEQ/Kg (5-10% of insulin producing islets normally used in intra-portaltransplants) provides a rigorous test of the cell transplantationdevices. Currently in clinical islet transplantation therapy, theinfusion of an adequate amount of β-cell mass has posed an obstacle fortreatment of insulin-dependent diabetes. Insulin independence isroutinely achieved when a sufficient quantity of islet cells aredelivered, approximately 10,000 IEQ/Kg of recipient's body weight. Toprovide this quantity of islet cells, present day islet transplantprotocols require more than one donor pancreas per recipient, creating astrain on an already limited donor supply. Therefore, if glycemiccontrol can be achieved using only 5-10% of the islets currently used inintra-portal transplants, the number of diabetic patients that couldreceive islet transplant therapy would increase significantly.

Histological analyses of explanted devices were performed to test thelong-term survival and function of transplanted islets. Islet graftfunction was also monitored through bi-weekly blood glucose andbi-monthly intravenous glucose tolerance tests (IVGTTs).

a) Histological Analysis of Islet Graft Function

Following explantation of the devices at 9-weeks, insulin was detectedin the devices using specific primary antibodies against insulin. FIG.17A shows the result of the insulin staining within the porous chamberof an explanted device. The detection of insulin within the chamberindicated that the islet cells contained in the device were viable andfunctional at 9-weeks post-transplantation. Immunohistochemical stainingof explant sections demonstrated healthy, well-configured isletssurrounded by robust microvessels (FIG. 17B; microvessels indicated byarrows).

b) Blood Glucose Measurements

Weekly fasting and non-fasting blood glucose levels were measured tomonitor for islet graft function following transplantation. Thesemeasurements aid in determining the overall efficacy of the celltransplantation devices in long-term control of blood glucose levels.Fasting blood glucose readings provide a controlled measure of graftfunction. Briefly, a drop (several microliters) of blood is collectedfrom a vein of a recipient animal, and the blood glucose level isdetermined using a Freestyle Lite glucometer or other glucose testingdevice.

As shown in FIG. 18 , the transplanted islets demonstrated long-termglucose control up to the explantation of the devices at 72 days. Theanimals in the “glycemic control” group (n=4) were insulin-independent,and the blood glucose levels were controlled by the islets in the celltransplantation devices alone. The animals in this group showedlong-term insulin independence after islet transplantation. Someanimals, however, remained hyperglycemic (elevated daily blood glucoselevels) following transplantation of islets into the devices (n=6). Thiswas related to poor metabolic quality of the pre-transplant islets andlow islet transplant dose (IEQ/Kg). The quality of islets prior totransplantation correlated well with long-term islet function.

c) Glucose Tolerance Test

Glucose tolerance tests are important in assessing islet graft functionthrough the comparison of pre- and post-transplant IVGTT results. Totest the efficacy of the cell transplantation devices, IVGTTs wereconducted prior to pancreatectomy (baseline), at various time pointsafter islet transplantation into the devices, and after explantation ofthe devices. IVGTT was performed by injecting a dose of dextrose andmeasuring the time it takes for endogenous insulin to bring the glucoselevels to baseline. In addition to measuring blood glucose level, bloodwas sampled at various time points to measure the level of C-peptide,which is a by-product created when insulin is produced by β cells.Results for an IVGTT were interpreted using absolute values of bloodglucose level (FIG. 19A), area under curve (AUC) of blood glucose level(FIG. 19B), and fold change in C-peptide level (FIG. 19C).

As shown in FIGS. 19A and 19B, the glucose levels rise significantly(p<0.001, Anova) after the device is explanted, indicating that theremoval of the device results in elimination of insulin function similarto a diabetic animal with no islets. While the lowest glucose levelswere detected in non-pancreatectomized animals, the islet autograftrecipients showed significant reduction is glucose levels after dextroseinjection, indicating that the immature islets can survive and functionafter transplantation.

Serum samples from the IVGTTs were analyzed using Linco's PorcineC-Peptide Radioimmunoassay kit, which utilizes an antibody madespecifically against synthetic porcine C-peptide. Serum samples at 0, 5,15, 30, 60 and 120 minutes post-dextrose injection were analyzed for thepresence of porcine C-peptide. Four study groups weretested—non-pancreatectomized pigs (baseline), islet autograft recipients(post-islet transplantation), autograft recipients that have had theirdevices removed (post-device removal) and diabetic control pigs. Whenexamining fold changes in C-peptide levels among the different studygroups, baseline and post-islet transplant recipients showed verycomparable result, although the C-peptide level in post-islet transplantrecipients increased at 60 minutes as opposed to 30 minutes for thebaseline group (FIG. 19C). Furthermore, fold changes in C-peptide forthe post-device removal group and diabetic control group were similar,indicating that the transplanted islets were responsible for C-peptiderelease prior to device removal.

Other embodiments of the invention will be apparent to those skilled inthe art from consideration of the specification and practice of theinvention disclosed herein. It is intended that the specification andexamples be considered as exemplary only, with a true scope and spiritof the invention being indicated by the following claims.

1. A device for implanting cells in a host body, comprising: a porousscaffold comprising at least one chamber having a proximal end and adistal end, the porous scaffold having pores sized to facilitate growthof vascular and connective tissues into the at least one chamber; and atleast one removable plug configured to be positioned within the at leastone chamber; wherein the porous scaffold comprises a polypropylene mesh.2-85. (canceled)